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FADD null mouse embryonic fibroblasts undergo apoptosis after photosensitization with the silicon phthalocyanine Pc 4
Abstract:
Oxidative stress, such as photodynamic therapy with the silicon phthalocyanine Pc 4 (Pc 4-PDT), can induce apoptosis and tumor necrosis factor alpha (TNF) production. TNF receptors, as well as other death receptors, have been implicated in stress-induced apoptosis. To assess directly the role of FADD, a death receptor-associated protein, in induction of apoptosis post-Pc 4-PDT, embryonic fibroblasts from FADD knock out (k/o) and wild-type (wt) mice were used. Pc 4-PDT induced casp-3 activation and apoptosis in both cell types. In the presence of zVAD, a pancaspase inhibitor, Pc 4-PDT-induced apoptosis was abrogated in both cell lines. Fumonisin B1 (FB), an inhibitor of ceramide synthase, had no effect on apoptosis after Pc 4-PDT in either cell line. Similar to Pc 4-PDT, exogenous C6-ceramide bypassed FADD deficiency and induced zVAD-sensitive apoptosis. In contrast to Pc 4 photosensitization, TNF did not induce either apoptosis or ceramide accumulation in FADD k/o cells. In the absence of FADD deficiency, TNF-induced apoptosis was zVAD-sensitive and FB-insensitive. Induced ceramide levels remained elevated after cotreatment with TNF and zVAD in FADD wt cells. Taken together, these data provide genetic evidence for a lack of FADD requirement in Pc 4-PDT- or C6-ceramide-induced apoptosis. FB-sensitive ceramide production accompanies, but does not suffice, for apoptosis after Pc 4 photosensitization or TNF.
Insights
Photodynamic therapy (PDT) induces apoptosis independently of FADD, a key protein in death receptor signaling. Ceramide production is involved but not sufficient for this cell death pathway.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Oxidative stress, induced by methods like silicon phthalocyanine Pc 4 photodynamic therapy (Pc 4-PDT), can trigger apoptosis and tumor necrosis factor alpha (TNF) production.
- TNF receptors and other death receptors are known to play roles in stress-induced apoptosis.
Purpose of the Study:
- To investigate the direct role of FADD (Fas-associated death domain), a protein associated with death receptors, in mediating apoptosis following Pc 4-PDT.
Main Methods:
- Utilized embryonic fibroblasts from FADD knockout (k/o) and wild-type (wt) mice.
- Assessed apoptosis and caspase-3 activation after Pc 4-PDT.
- Employed pancaspase inhibitor zVAD and ceramide synthase inhibitor Fumonisin B1 (FB).
- Administered exogenous C6-ceramide and TNF to evaluate their effects in FADD-deficient cells.
Main Results:
- Pc 4-PDT induced caspase-3 activation and apoptosis in both FADD k/o and wt cells.
- Apoptosis induced by Pc 4-PDT was blocked by zVAD in both cell types.
- Fumonisin B1 (FB) did not affect Pc 4-PDT-induced apoptosis.
- Exogenous C6-ceramide induced apoptosis independently of FADD, sensitive to zVAD.
- TNF failed to induce apoptosis or ceramide accumulation in FADD k/o cells.
- TNF-induced apoptosis in wt cells was zVAD-sensitive and FB-insensitive.
Conclusions:
- These findings provide genetic evidence that FADD is not required for apoptosis induced by Pc 4-PDT or C6-ceramide.
- FB-sensitive ceramide production occurs during Pc 4-PDT and TNF-induced apoptosis but is not sufficient to cause it.